Aims: To determine the effects of water activity (a(W); 0 center dot 995-0 center dot 90), temperature (5, 18, 25 and 30 degrees C), time of incubation (7-35 days) and their interactions on tenuazonic acid (TA) production on 2% soybean-based agar by two Alternaria alternata strains isolated from soybean in Argentina. Methods and Results: TA production by two isolates of A. alternata was examined under interacting conditions of a(W), temperature and time of incubation on 2% soybean-based agar. Maximum TA production was obtained for both strains at 0 center dot 98 a(W), but at 30 and 25 degrees C for the strains for RC 21and RC 39, respectively. The toxin concentration varied considerably depending on a(W), temperature, incubation time and strain interactions. TA was produced over the temperature range from 5 to 30 degrees C and a(W) range from 0 center dot 92 to 0 center dot 995, however at 5 and 18 degrees C little TA was produced at a(W) below 0 center dot 94. Contour maps were developed from these data to identify areas where conditions indicate a significant risk for TA accumulation. Conclusions: The optimum and marginal conditions for TA production by A. alternata on soybean-based agar were identified. The results indicated that TA production by A. alternata is favoured by different temperatures in different strains. Significance and Impact of the Study: Data obtained provide very useful information for predicting the possible risk factors for TA contamination of soybean as the a(W) and temperature range used in this study simulate those occurring during grain ripening. The knowledge of TA production under marginal or sub-optimal temperature and a(W) conditions for growth are relevant as improper storage conditions accompanied by elevated temperature and moisture content in the grain can favour further mycotoxin production and lead to reduction in grain quality.